Circulating antibodies determination

The age of the recipient is another important determining factor in vaccine and toxoid response. In the first few months of life, maternal antibodies acquired via transplacental transfer during the third trimester of gestation protect an infant. However, the maternal antibody also inhibits the immune response to live vaccines because the circulating antibodies neutralize the vaccine before the infant has the opportunity to mount an immune response. For this reason, live vaccines are not administered until maternal antibody has waned, generally by age 12 months. ... 

Determination of the cellular source of intrinsic factor followed introduction of radioactive vitamin B12 and the discovery in the late 1950s of circulating antibodies to components of the gastric mucosa. 

Lymphocytes, the effector cells of the acquired immune system, include morphologically indistinguishable T and B cells, the former divided into CD4+ T helper cells and CD8+ cytotoxic T cells. Since the functions of those cell subsets differ so drastically, it became important to develop tools to distinguish them from each other. Efforts to identify cell subsets according to their expression of different surface antigens have been successful, including various Cluster of Determination (CD) markers (Table 23.1). In addition, cross-reactive monoclonal antibodies, and subsequently developed species-specific polyclonal and monoclonal antibodies towards the major histocompatibility complex (MHC) have been used to label cells in circulation and in tissue sections (Table 23.1). 

Measurement of the serum concentrations of administered antibodies is a general tool to evaluate their persistence in circulation. This is usually performed by introducing a sufficient amount of the test antibody either by the intravenous or by the intraperi-toneal routes (see Note 3), in a quantity that can be easily detected and quantified in serum samples, even after a two log reduction in concentration. The antibody tracer can be labeled with radioisotope which permits direct quantification in serum samples. To minimize radioactive isotope use, we use an antibody tracer that is unmodified or labeled with biotin or other derivation chemistries, and then determine its serum concentrations by ELISA techniques. We commonly inject 100 xg of the test antibody in a 200 xl volume of phosphate-buffered saline into each mouse intraperitoneally (i.p.) (see Note 4). This amount can vary depending on the goals of the experiment and the sensitivity of the detection method. A minimum of five inbred mice, sex-matched and age-matched, at 8-16 weeks of age are recommended for each antibody to be tested. 

Pharmacokinetic data were collected as well as pharmacodynamic measurements of platelet aggregation support (ristocetin cofactor activity) and cuticle wound blood flow. An important component of these studies was the suitability of the model. These models were chosen because of the biochemical deficiency of the particular factors and the parallel clinical syndromes. Such in vivo data can help in determining activity and dosing when such a product is first used in human trials. The Refacto molecule was also studied in rats and monkeys to determine its no observed adverse effect level, that was more than 10 times normal circulating levels. The major toxicity observed was the development of antibodies to the molecule that blocked activity and resulted in an acquired hemophilia syndrome. Similar findings were demonstrated when plasma-derived material was injected into monkeys [20]. 

The determination of circulating anti-insulin antibodies is of clinical importance for the following reasons ... 

Preprothrombin can be determined immunologically, either using antiprothrombin antibodies, after adsorption of the y -carboxylated protein onto barium carbonate or using anti-preprothrombin antibodies that do not cross-react with prothrombin. Circulating concentrations of preprothrombin in vitamin K deficiency are of the order of 150 to 1,500 nmol per L. If elevated preprothrombin is because of vitamin K deficiency, then it will fall on administration of the vitamin, whereas if it is the result of liver disease, then vitamin K supplements will have no effect. 

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